Field of Invention
The invention relates to systems and methods for producing bio-materials, such as pharmaceutical proteins.
Description of Related Art
A photoautotrophic microorganism is an organism that is capable of generating its own sustenance from inorganic substances using light as an energy source. As an example, photosynthetic microscopic algae, hereinafter referred to as algae, is a photoautotroph. Algae are unicellular organisms which produce oxygen by photosynthesis, and may include flagellates, diatoms, and blue-green algae. More than 100,000 species of algae are known, and it is expected that the actual number of species exceeds 2,000,000. This count does not include genetically modified algae species.
As mentioned above, algae use a photosynthetic process similar to that of higher-developed plants, with certain advantages not found in traditional crops, such as rapeseed, wheat, or corn. Algae have a high growth rate, and it is possible to complete an entire harvest in hours. Further, algae are tolerant to varying environmental conditions, for example, algae may be grown in saline waters that are unsuitable for agriculture. Due to this tolerance to environment and climate, algae are responsible for about one-third of the net photosynthetic activity worldwide.
As a result, during the past decade, much focus has been aimed at the production of algae for commercial purposes. This focus is evidenced by the manifestation of many new industries and uses of algal production, including but not limited to the following: (i) use of algae as a source of fatty acids, proteins and other bio-chemicals in the production of nutraceuticals, health food, food additives, vitamins, pharmaceuticals, and natural dyes; (ii) use of algae as an animal feed supplement with nutritional value equivalent to that of soybean meal; (iii) use of algae as a biological control of agricultural pests; (iv), use of algae as soil conditioners and bio-fertilizers in agriculture; (v) use of algae for the production of oxygen and removal of nitrogen, phosphorus, and toxic substances in sewage treatment; (vi) use of algae in the bio-degradation of plastics; (vii) use of algae as a renewable biomass source for the production of a diesel fuel substitute (biodiesel) and other biofuels such as ethanol, methane gas, and hydrogen; and (viii) use of algae to scrub CO2, NOx, VOx from effluent released during the production of fossil fuel. With so many uses, it would be desirable to mass produce algae in a low-cost, high-yield manner.
One commercial purpose of significant import includes algae production as a renewable biomass source for sustainable biodiesel production. Presently, the renewable biomass source is provided by edible oils, such as soybean oil, palm oil, and rapeseed oil. It is of interest to develop additional types of renewable biomass sources, such as lipids from algae.
As briefly noted above, one factor making algae interesting as a renewable biomass source is that algae may be grown under conditions or in places not suitable to other sources. Accordingly, algae may be grown and used in ways that do not significantly compete with food sources or agriculturally productive land.
An additional factor making algae of interest is the availability of material to convert to biodiesel or other fuel. Some algae have lipid content as much as 50% to 70% of their dry weight. By way of comparison, the lipid content in dry soybeans is approximately 20%. Algal lipids have a similar composition to vegetable oil and are readily adaptable as a renewable biomass source to existing biodiesel manufacturing processes. Further, the remaining algal biomass may be converted to bio-ethanol, converted to biodiesel, converted to methane, burned, or used as food for other organisms.
As an additional interest, algae can be exploited as commercial sources of Omega 3, Omega 6 oils and Astaxanthin as precursors for pharmaceuticals, nutraceuticals and food supplements.
Commercial acceptance of biomass products is dependent on a variety of factors such as, for example, cost to manufacture, cost to operate, reliability, durability, and scalability. Commercial acceptance of biomass products is also dependent on the ability to increase biomass product growth and recovery, while decreasing biomass production cost. Therefore, it may be desirable to have novel approaches for growing and harvesting biomass products including, for example, cell components such as lipids, proteins, vitamins, fatty acids, minerals, carotenoids, pigments, and the like.
Providing faster growth and producing high density cultures is critical to achieving the operational scale necessary for current environmental and industrial needs. Ideally, improving the speed of growth and increasing the density of a culture will require less production space and consequently will lower the cost of associated facilities.
In line with these reasons and others, the cultivation of algae in liquid suspension compared to algae in stagnant pools allows greater access to the nutrient sources necessary for growth, i.e., water, CO2, and minerals, and permits reducing the production space to a cost-effective footprint. Accordingly, considerable activity has been focused on efficiently growing photoautotrophic microorganisms in liquid suspension, and specifically to mass culture unicellular algae.
Algae production yields are currently adversely affected by many factors. Algal yield can be restricted by the limited wavelength range of light energy capable of driving photosynthesis, between about 400-700 nm (nanometers), which is only about half of the total solar energy. Other factors, such as respiration requirements during dark periods, efficiency of absorbing sunlight, and other growth conditions can affect photosynthetic efficiencies in algal bioreactors. The net result is an overall photosynthetic efficiency that has been too low for economical large scale production. Thus, the need exists for a large scale production system that provides the user a cost-effective means of installation, operation and maintenance relative to production yields. It is desirable that such a system also increase photosynthesis to maximize production yield.
In order to produce optimal yields, algae need to have CO2 in large quantities in the basins or bioreactors where they grow. In addition to CO2, the growth rate of algae may benefit from exposure to other nutrients that are common in known plant fertilizers.
Furthermore, algae need effective control of light. To maximize the growth of photosynthetic organisms, light must be available at the right intensity, the right wavelength, the right frequency, and without excessive heat. Excessive light intensity can limit growth by inducing photo-respiration or bleaching the pigments needed for efficient cell growth. In addition, light intensity or light frequency in excess of the culture requirements may result in heat build-up that can limit culture growth. These problems are readily apparent in production systems that rely solely on direct solar light as a driver of photosynthesis, such as in ponds and raceways. Solar light is subject to extreme diurnal and seasonal variability along with local weather conditions. In addition, cultures relying on direct solar light are subject to periodic heating from light intensities and spectra not immediately useable by the culture.
At high growth rates, algae biomass production is limited by three factors, oxygen content, growth byproducts, and light intensity. Dissolved oxygen becomes a hindrance that limits growth and can poison the algae in the media. Removal of dissolved oxygen is most often accomplished by bubbling air through the media to exchange dissolved oxygen existing above equilibrium with the 20% level in found in air.
During algae production cycles, the media becomes filled with growth byproducts. Richmond, et al described how these byproducts limit growth. Byproducts are often removed by bubbling and subsequently removing the bubbles from the process. Algae growth byproducts are varied but contain oils and other chemicals that readily form bubbles. The bubbles are forced through a small aperture then rinsed to destroy the bubble structure and allow more room for the next bubbles while collecting byproducts and removing them from the media.
To grow at high rates, it is important to efficiently illuminate bio-material, such as algae, throughout the fluid medium using high light intensity. However, high light intensity causes changes in the algae usually referred to as photo-inhibition. Photo-inhibition is characterized by several processes. One such process is the growth of carotenoids that shield the chlorophyll from the light and reduce production. The process has evolved to protect algae that usually operate naturally at low light levels ˜70 micromoles/m^2/sec (one sun is 2000 micromole/^2/sec). The second process is a shortening of the antenna between generations of algae in response to intense light. To grow in intense light efficiently, two conditions must be met: (1) the algae must be at high concentration (gm/liter); and (2) the algae must be moved quickly between light and dark regions of the media so that on average the algae get the low level light. The time for these bright flashes and periods of dark are important and must occur on millisecond time scales.
As an example, it is desirable to grow Spirulina at high rates at high density with high light intensity. The algae are circulated to produce light and dark pulsation of the light irradiation using almost violent bubbling of the densely loaded media. This method is common and appropriate for Spirulina, which is one of the most damage resistant cyanobacteria or algae species known. Other algae species would be damaged by the mechanical damage and shear force in the media flow pattern.
Pumping of algae is common in many algae growth systems. The most common method of pumping is via air lift. Air lift pumping is always counter gravity because it is caused by a difference in density. Air lifting is very energy inefficient and limited on the pumping rate by shear forces that occur at several phases of the pumping (reference). Air lift is best for high flow rate at very low head pressure.
Pumping is a means of damaging algae and providing non-viable cells in the media. Viable algae cells have defenses against attack by bacterial and virus, but non viable cells become food for bacterial and virus. One aim of the development is to provide an environment where algae has high growth rate advantages over bacteria and virus. This environment will enable long time between inoculation and increase production efficiency.
Any media requires fundamental parameters to be productive. The media must have correct nutrients for efficient growth of the algae. For example, sufficient carbon, usually in the form of CO2, must be present. The temperature must be optimized for growth rate. Light must be supplied at levels that match the ability of algae to absorb it efficiently and convert it efficiently to algae biomass. High intensity changes the algae sensitivity to light resulting in a mixed optimum light intensity.
The ability to effectively control light for 24 hours per day as well as continuous consumption of CO2 and other nutrients, encourages faster growth of biomass and secondary metabolites.
These factors result in a more efficient use of facilities, enabling a smaller footprint for a given level of production. However, the use of artificial light has a cost that must be minimized for successful industrial application.
The two major obstacles that reduce the uniform delivery of light to a culture are turbidity and bio-filming. Turbidity occurs as a culture approaches a density where some of the organisms shade others from the light. Ensuring delivery of the appropriate amount of light to each organism becomes increasingly difficult as the density of the organisms in a culture increases. Turbidity within a culture results in some organisms receiving less light than they can utilize while other organisms receive non-productive, or even harmful absorption of light. This absorption of excess light wastes energy and contributes to heat build-up.
Bio-filming is an extremely widespread problem that occurs when a microorganism adheres to a surface. Most microorganisms, including species in all three domains, i.e., bacteria, eukaryotes, and archaea, perform processes that result in adhesion to surfaces and to other microorganisms. In industrial applications, biofilms often clog or corrode pipes and surfaces. In photo-bioreactors a biofilm can form over a light-delivery surface, thereby reducing the intensity and changing the spectra of the light transmitted.